Not Applicable
Not Applicable
Degradation of extracellular matrix (ECM) is critical for normal embryonic development, blastocyst implantation, organ morphogenesis, nerve growth, ovulation, cervical dilatation, postpartum uterine involution, endometrial cycling, hair follicle cycling, bone remodeling, wound healing, angiogenesis, apoptosis, and numerous other normal physiological processes. Aberrant degradation of ECM is associated with numerous disease states including, for example, tissue invasion and metastasis by tumor cells, aberrant angiogenesis, cardiovascular diseases (e.g., heart failure and atherosclerosis), arthritis, nephritis, neurological diseases (e.g., macular degeneration), breakdown of the blood-brain barrier, periodontal disease, skin ulceration, gastric ulceration, corneal ulceration, liver fibrosis, emphysema, fibrotic lung disease, and other pathological conditions.
Secreted proteinases of the extracellular matrix metalloproteinase (MMP) gene family are essential for maintaining the architecture of tissues and organs. MMPs are sometimes designated xe2x80x98matrixins,xe2x80x99 and catalyze degradation of ECM. The protein degradative activity of MMPs can be modulated by endogenous inhibitors, alpha-macroglobulins, and tissue inhibitors of metalloproteinases (TIMPs). In addition, expression of most MMPs is transcriptionally regulated by one or more growth factors, hormones, and cytokines, and tends to be differentially regulated during various phases of cellular transformation.
MMPs are generally synthesized in the form of catalytically inactive pre-pro-enzymes, and secreted in the form of inactive pro-enzymes. Pro-MMPs can be activated in vitro using various proteases, thiol-reactive agents, mercurial compounds, reactive oxygen, and denaturing agents. In vivo, pro-MMPs are believed to be activated by removal of the propeptide domain, catalyzed by tissue or plasma proteinases, or sometimes by opportunistic bacterial proteinases. In vivo activation of pro-MMPs is believed to occur primarily at cell surfaces.
Although a primary function of MMPs is catalysis of simple removal (i.e., resorption) of ECM, MMPs can also alter the biological activity exhibited by ECM components. For example, MMP-2 released from neurite growth cones inactivates neurite growth-inhibiting chondroitin sulfate proteoglycans and uncovers laminin (which enhances neurite growth). Other normal and pathological processes in which MMP-catalyzed changes in ECM protein structures have been implicated are described, for example in Nagase et al. (1999, J. Biol. Chem. 274:21491-21494).
No fewer than 23 classes of MMPs, characterized by no fewer than eight characteristic domain arrangements, have been described. In view of the widespread and critical nature of MMP activities in normal and pathological physiological processes, a need exists for identification of further members of this protein family. The present invention satisfies this need by providing a novel human MMP.
The present invention is based, in part, on the discovery of a novel gene encoding an MMP, the gene being referred to herein as xe2x80x9c46798xe2x80x9d. cDNAs encoding 46798 have been isolated in at least two different forms, herein designated the xe2x80x98shortxe2x80x99 and xe2x80x98longxe2x80x99 forms. The nucleotide sequence of a cDNA encoding the long form of 46798 is shown in SEQ ID NO: 1, and the amino acid sequence of the long form of the 46798 polypeptide is shown in SEQ ID NO: 2. In addition, the nucleotide sequence of the coding region is depicted in SEQ ID NO: 3. The nucleotide sequence of a cDNA encoding the short form of 46798 is shown in SEQ ID NO: 11, and the amino acid sequence of the short form of the 46798 polypeptide is shown in SEQ ID NO: 12. In addition, the nucleotide sequence of the coding region is depicted in SEQ ID NO: 13. The short and long forms of 46798 are individually and collectively referred to herein as xe2x80x9846798 proteinsxe2x80x99 or xe2x80x9846798 nucleic acids.xe2x80x99
Accordingly, in one aspect, the invention features a nucleic acid molecule that encodes a 46798 protein or polypeptide, e.g., a biologically active portion of the 46798 protein. In a preferred embodiment the isolated nucleic acid molecule encodes a polypeptide having the amino acid sequence of either of SEQ ID NOs: 2 and 12. In other embodiments, the invention provides isolated 46798 nucleic acid molecules having the nucleotide sequence of one of SEQ ID NOs: 1, 3, 11, and 13. In still other embodiments, the invention provides nucleic acid molecules that have sequences that are substantially identical (e.g., naturally occurring allelic variants) to the nucleotide sequence of one of SEQ ID NOs: 1, 3, 11, and 13. In other embodiments, the invention provides a nucleic acid molecule which hybridizes under stringent hybridization conditions with a nucleic acid molecule having a sequence comprising the nucleotide sequence of one of SEQ ID NOs: 1, 3, 11, and 13, wherein the nucleic acid encodes a full length 46798 protein or an active fragment thereof.
In a related aspect, the invention further provides nucleic acid constructs that include a 46798 nucleic acid molecule described herein. In certain embodiments, the nucleic acid molecules of the invention are operatively linked to native or heterologous regulatory sequences. Also included are vectors and host cells containing the 46798 nucleic acid molecules of the invention, e.g., vectors and host cells suitable for producing 46798 nucleic acid molecules and polypeptides.
In another related aspect, the invention provides nucleic acid fragments suitable as primers or hybridization probes for detection of 46798-encoding nucleic acids.
In still another related aspect, isolated nucleic acid molecules that are antisense to a 46798-encoding nucleic acid molecule are provided.
In another aspect, the invention features 46798 polypeptides, and biologically active or antigenic fragments thereof that are useful, e.g., as reagents or targets in assays applicable to treatment and diagnosis of 46798-mediated or related disorders (e.g., MMP-mediated disorders such as those described herein). In another embodiment, the invention provides 46798 polypeptides having matrix metalloproteinase activity. Preferred polypeptides are 46798 proteins including at least one MMP domain, and preferably having a 46798 activity, e.g., a 46798 activity as described herein. Preferred polypeptides are 46798 proteins including at least one transmembrane domain and at least one peptidase_M10 domain.
In other embodiments, the invention provides 46798 polypeptides, e.g., a 46798 polypeptide having the amino acid sequence shown in one of SEQ ID NOs: 2 and 12; an amino acid sequence that is substantially identical to the amino acid sequence shown in one of SEQ ID NOs: 2 and 12; or an amino acid sequence encoded by a nucleic acid molecule having a nucleotide sequence which hybridizes under stringent hybridization conditions to a nucleic acid molecule comprising the nucleotide sequence of any of SEQ ID NOs: 1, 3, 11, and 13, wherein the nucleic acid encodes a full length 46798 protein or an active fragment thereof.
In a related aspect, the invention further provides nucleic acid constructs that include a 46798 nucleic acid molecule described herein.
In a related aspect, the invention provides 46798 polypeptides or fragments operatively linked to non-46798 polypeptides to form fusion proteins.
In another aspect, the invention features antibodies and antigen-binding fragments thereof, that react with, or more preferably, specifically bind, 46798 polypeptides.
In another aspect, the invention provides methods of screening for compounds that modulate the expression or activity of the 46798 polypeptides or nucleic acids.
In still another aspect, the invention provides a process for modulating 46798 polypeptide or nucleic acid expression or activity, e.g., using the screened compounds. In certain embodiments, the methods involve treatment of conditions related to aberrant activity or expression of the 46798 polypeptides or nucleic acids, such as conditions involving aberrant or deficient degradation or resorption of ECM proteins or aberrant or deficient proteolytic activation of extracellular matrix proteins.
The invention also provides assays for determining the activity of or the presence or absence of 46798 polypeptides or nucleic acid molecules in a biological sample, including for disease diagnosis.
In further aspect the invention provides assays for determining the presence or absence of a genetic alteration in a 46798 polypeptide or nucleic acid molecule, including for disease diagnosis.
Other features and advantages of the invention will be apparent from the following detailed description, and from the claims.